The world needs a highly effective malaria vaccine. Sanaria(r) PfSPZ Vaccine, composed of aseptic, purified, cryopreserved radiation attenuated PfSPZ, protected 100% of volunteers in an NIH clinical trial. The vaccine has been administered by direct venous inoculation (DVI) to 371 subjects in Mali, Tanzania, Equatorial Guinea (EG), and USA. Safety, tolerability, high-grade protection, heterologous protection after Controlled Human Malaria Infection, durable protection in the field, efficacy with 3 doses and safety at 5x the 100% protective dose, have been established. Sanaria has met with the World Health Organization to plan for pre-qualification and establishment of a Technical Advisory Group for PfSPZ Vaccine administered by DVI. In 2015-16, the vaccine will be assessed by DVI in Tanzania, Kenya (500 infants), Mali, Burkina Faso, EG, Germany, and USA. The first licensure submission in USA is planned for late 2017. Reduction in the number of PfSPZ/ regimen, number of doses, and/or time to complete an immunization regimen, and prolongation of efficacy will reduce costs and improve implementation. Being able to immunize by cutaneous or intramuscular routes will increase capacity to immunize young infants and facilitate immunization by less experienced personnel. To these ends, we have studied adjuvants that augment/prolong immune responses. Traditional and experimental adjuvants, including multiple toll-like receptor ligands, do not work, likely because the vaccine is composed of live-attenuated organisms and no licensed adjuvants are known to enhance the CD8+ T cell-mediated immunity which appears to underlie protective efficacy against PfSPZ Vaccine. A novel glycolipid, 7DW8-5, which binds CD1d and stimulates iNKT cells, has strong adjuvant effects in mice immunized with irradiated P. yoelii sporozoites (irrPySPZ), and enabled reduction to one dose when irrPySPZ were administered by DVI (75% protection) and from 4 to 2 doses when administered intradermally (70%-100% protection). Protection lasted for at least 14 weeks. Four DVI doses during a week of 2x103 purified, cryopreserved irrPySPZ with and without 7DW8-5 protected 96% (15/16) and 44% (7/16) of mice respectively (p=0.0059), raising the possibility of an accelerated immunization regimen that would be ideal for travelers and mass administration campaigns. In non-human primates (NHPs), 7DW8-5 with PfSPZ Vaccine was well tolerated; greatly enhancing the magnitude of splenic CD8+ and CD4+ T cell responses 2.5 months post vaccination. These findings support development of 7DW8-5 for use with PfSPZ Vaccine administered both by intravascular and traditional routes to allow for reduced cost of goods, rapid immunization and increased durability of protection. In this project we will assess 7DW8-5 with rodent, human, and simian SPZ in mice and NHPs, and manufacture 7DW8-5 in compliance with cGMPs. Accomplishing the Specific Aims of this proposal will provide the foundation for the first phase 1 clinical trial of PfSPZ Vaccine administered with 7DW8-5, and eventual use of this combination for rapid and cost effective immunization of travelers/military and for mass malaria elimination campaigns.